Interferon-alpha 54

ABSTRACT

A new polypeptide, called IFN-α54, produced by E. coli transformed with a newly isolated and characterized human IFN-α gene is described. The polypeptide exhibits interferon activities such as antiviral activity, cell growth regulation, and regulation of production of cell-produced substances.

CROSS-REFERENCE TO RELATED APPLICATION

This application is a continuation-in-part of copending U.S. Ser. No339,828 filed 15 Jan. 1982, now abandoned.

DESCRIPTION

1. Technical Field

The invention is in the field of biotechnology. More particularly itrelates to a polypeptide having interferon (IFN) activity, DNA thatcodes for the polypeptide, a recombinant vector that includes the DNA, ahost organism transformed with the recombinant vector that produces thepolypeptide, pharmaceutical compositions containing the polypeptide, andtherapeutic methods employing the polypeptide.

2. Background Art

IFNs are proteins with antiviral, immunomodulatory, andantiproliferative activities produced by mammalian cells in response toa variety of inducers (see Stewart, W. E., The Interferon System,Springer-Verlag, New York, 1979). The activity of IFN is largely speciesspecific (Colby, C., and Morgan, M. J., Ann. Rev. Microbiol. 25:333-360(1971) and thus only human IFN can be used for human clinical studies.Human IFNs are classified into three groups, α, β, and γ, (Nature,286:110, (1980)). The human IFN-α genes compose a multigene familysharing 85%-95% sequence homology (Goeddel, D. V. et al, Nature290:20-27 (1981) Nagata, S., et al, J. Interferon Research 1:333-336(1981)). Several of the IFN-α genes have been cloned and expressed in E.coli (Nagata, S., et al, Nature 284:316-320 (1980); Goeddel, D. V., etal, Nature 287:411-415 (1980); Yelverton, E., et al, Nucleic AcidsResearch, 9:731-741, (1981); Streuli, M., et al, Proc Nat Acad Sci(USA), 78:2848-2852. The resulting polypeptides have been purified andtested for biological activities associated with partially purifiednative human IFNs and found to possess similar activities. Accordinglysuch polypeptides are potentially useful as antiviral, immunomodulatory,or antiproliferative agents.

A principal object of the present invention is to provide a polypeptidehaving interferon activity that is produced by an organism transformedwith a newly isolated and newly characterized IFN-α gene. Thispolypeptide is sometimes referred to herein as IFN-αlb 54. Other objectsof the invention are directed to providing the compositions and hoststhat are used to produce this polypeptide and to therapeuticcompositions and methods that use this polypeptide as an activeingredient.

DISCLOSURE OF THE INVENTION

One aspect of the invention is a polypeptide having interferon activityand comprising the amino acid sequence:

    ______________________________________                                        CysAspLeuProGln    ThrHisSerLeuGly                                            HisArgArgThrMet    MetLeuLeuAlaGln                                            MetArgArgIleSer    LeuPheSerCysLeu                                            LysAspArgHisAsp    PheArgPheProGln                                            GluGluPheAspGly    AsnGlnPheGlnLys                                            AlaGluAlaIleSer    ValLeuHisGluVal                                            IleGlnGlnThrPhe    AsnLeuPheSerThr                                            LysAspSerSerVal    AlaTrpAspGluArg                                            LeuLeuAspLysLeu    TyrThrGluLeuTyr                                            GlnGlnLeuAsnAsp    LeuGluAlaCysVal                                            MetGlnGluValTrp    ValGlyGlyThrPro                                            LeuMetAsnGluAsp    SerIleLeuAlaVal                                            ArgLysTyrPheGln    ArgIleThrLeuTyr                                            LeuThrGluLysLys    TyrSerProCysAla                                            TrpGluValValArg    AlaGluIleMetArg                                            SerPheSerSerSer    ArgAsnLeuGlnGlu                                            ArgLeuArgArgLys    Glu                                                        ______________________________________                                    

A second aspect of the invention is a DNA unit or fragment comprising anucleotide sequence that encodes the above described polypeptide.

A third aspect of the invention is a cloning vehicle or vector thatincludes the above described DNA.

A fourth aspect of the invention is a host that is transformed with theabove described cloning vehicle and that produces the above describedpolypeptide.

A fifth aspect of the invention is a process for producing the abovedescribed polypeptide comprising cultivating said transformed host andcollecting the polypeptide from the resulting culture.

Another aspect of the invention is a pharmaceutical composition havinginterferon activity comprising an effective amount of the abovedescribed polypeptide admixed with a pharmaceutically acceptablecarrier.

Still another aspect of the invention is a method of providinginterferon therapy to a human comprising administering a therapeuticallyeffective amount of the above described polypeptide to the human.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a partial restriction map which shows the two XhoIIrestriction sites that produce a homologous 260 base pair DNA fragmentfrom the IFN-α1 and IFN-α2 structural genes. Data for this map are fromStreuli, M., et al Science, 209:1343-1347 (1980).

FIG. 2 depicts the sequencing strategy used to obtain the complete DNAsequence of the IFN-α54 gene coding region. Bacteriophage mp7:α54-1 DNAserved as the template for sequences obtained with primers A, H and Fand bacteriophage mp7:α54-2 DNA was the template for sequences obtainedwith primers E and G. The crosshatched area of the gene depicts theregion that encodes the 23 amino acid signal peptide and the open boxdepicts the region that encodes the mature polypeptide. The scale, inbase pairs, is numbered with 0 representing the ATG start codon ofpreinterferon. The arrows indicate the direction and extent ofsequencing with each primer.

FIG. 3 is the nucleotide sequence of the structural gene coding forIFN-α54 including some of the flanking 5'- and 3'- noncoding regions ofthe gene. The region coding for preinterferon and the mature polypeptidebegins with the ATG codon at position 61 and terminates with theTAA-codon at position 628.

FIG. 4 is a partial restriction map of the coding region of the IFN-α54gene. The crosshatching represents the region that encodes the 23 aminoacid signal peptide and the open box represents the gene coding sequencefor the mature polypeptide. The scale, in base pairs, is numbered with 0representing the ATG start codon of preinterferon.

FIG. 5 shows the amino acid sequence of the 23 amino acid signalpolypeptide and the 166 amino acid mature IFN-α54 coded for by the genedepicted in FIG. 3. The 189 amino acid sequence is displayed above thecorresponding nucleotide sequence. Amino acid 24, cysteine, is the firstamino acid of the mature IFN-α54 protein.

FIG. 6 is the DNA sequence of the E. coli trp promoter and the gene ofFIG. 3 which was inserted between the EcoRI and PvuII sites of theplasmid pBWll. The amino acid sequence of FIG. 5 is written above thecorresponding DNA sequence and the location of the restriction sitesused in the construction of the expression plasmid are indicated.

FIG. 7 is a diagram of the expression plasmid, pCS12.

MODES FOR CARRYING OUT THE INVENTION

In general terms IFN-α54 was made by identifying and isolating theIFN-α54 gene by screening a library of human genomic DNA with anappropriate IFN-α DNA probe, constructing a vector containing theIFN-α54 gene, transforming microorganisms with the vector, cultivatingtransformants that express IFN-α54 and collecting IFN-α54 from theculture. A preferred embodiment of this procedure is described below.

DNA PROBE PREPARATION

Total cytoplasmic RNA was extracted from human lymphoblastoid cells,Namalwa, which had been induced for IFN production by pretreatment with5-bromodeoxyuridine and Newcastle Disease Virus (NDV). The poly(A)(polyadenylic acid)-containing messenger RNA (mRNA) was isolated fromtotal RNA by chromatography on oligo(dT)-cellulose (type 3 fromCollaborative Research; Aviv, H., and Leder, P., Proc Natl Acad Sci(USA), 69:1408-1412, (1972)) and enriched for IFN mRNA by densitygradient centrifugation on 5%-20% sucrose gradients. Fractionscontaining IFN mRNA were identified by translating the mRNA bymicroinjecting aliquots of each fraction into Xenopus oocytes anddetermining the IFN activity of the products of the translationsaccording to a method described by Colman, A., and Morser, J., Cell,17:517-526 (1979).

The Namalwa cell human IFN enriched mRNA was used to constructcomplementary DNA (cDNA) clones in E. coli by the G/C tailing methodusing the PstI site of the cloning vector pBR322 (Bolivar, F., et al,Gene, 2:95-113 (1977)). A population of transformants containingapproximately 50,000 individual cDNA clones was grown in one liter ofmedium overnight and the total plasmid DNA was isolated therefrom.

The sequences of two IFN-α clones (IFN-α1 and IFN-α2) have beenpublished (Streuli, M., et al, Science, 209:1343-1347 (1980)).Examination of the DNA sequences of these two clones revealed that therestriction enzyme XhoII would excise a 260 bp fragment from either theIFN-α1 or the IFN-α2 gene (see FIG. 1). XhoII was prepared in accordancewith the process described by Gingeras, T. R., and Roberts, R. J., J MolBiol, 118:113-122 (1978).

One mg of the purified total plasmid DNA preparation was digested withXhoII and the DNA fragments were separated on a preparative 6%polyacrylamide gel. DNA from the region of the gel corresponding to 260bp was recovered by electroelution and recloned by ligation into theBamHI site of the single strand bacteriophage m13:mp7. Thirty-six cloneswere picked at random and the single stranded DNA isolated therefrom andsequenced. The DNA sequences of four of these clones were homologous toknown IFN-α DNA sequences. Clone mp7:α-260, with a DNA sequenceidentical to IFN-α1 DNA (Streuli, M. et al, Science, 209:1343-1347(1980)) was chosen as a highly specific hybridization probe foridentifying additional IFN-α DNA sequences. This clone is hereinafterreferred to as the "260 probe."

Screening of Genomic DNA Library

In order to isolate other IFN-α gene sequences, a ³² P-labelled 260probe was used to screen a library of human genomic DNA by in situhybridization. The human gene bank, prepared by Lawn, R. M., et al,Cell, 15:1157-1174 (1978), was generated by partial cleavage of fetalhuman DNA with HaeIII and AluI and cloned into bacteriophage λ Charon 4Awith synthetic EcoRI linkers. Approximately 800,000 clones werescreened, of which about 160 hybridized with the 260 probe. Each of the160 clones was further characterized by restriction enzyme mapping andcomparison with the published restriction maps of 10 chromosomal IFNgenes (Nagata, S., et al, J Interferon Research, 1:333-336 (1981)). Oneof the clones, hybrid phage λ4A:α54 containing a 16 kb insert, wascharacterized as follows. A DNA preparation of λ4A:α54 was cleaved withHindIII, BglII, and EcoRI respectively, the fragments separated on anagarose gel, transferred to a nitrocellulose filter (Southern, E. M., JMol Biol 98:503-517 (1977)) and hybridized with ³² P-labelled 260 probe.This procedure localized the IFN-α54 gene to a 3.9 kb EcoRI restrictionfragment which was then isolated and recloned, in both orientations, byligation of the fragment into EcoRI cleaved m13:mp7. The two subclonesare designated mp7:α54-1 and mp7:α54-2. The -1 designation indicatesthat the single-stranded bacteriophage contains insert DNA complementaryto the mRNA (the minus strand) and the -2 designation indicates that theinsert DNA is the same sequence as the mRNA (the plus strand).

Sequencing of the IFN-α54 Gene

The Sanger dideoxy-technique was used to determine the DNA sequence ofthe IFN-α54 gene. The strategy employed is diagrammed in FIG. 2, the DNAsequence thus obtained is given in FIG. 3, and a partial restrictionenzyme map of the IFN-α54 gene is illustrated in FIG. 4. Unlike manygenes from eukaryotic organisms, but analogous to other IFN chromosomalgenes which have been characterised 10 (Nagata, S., et al, Nature,287:401-408 (1980); Lawn, R. M., et al, Science, 212:1159-1162 (1981);Lawn, R. M., et al, Nucleic Acids Res., 9:1045-1052 (1981); Nagata, S.,et al, J Interferon Research, 1:333-336 (1981); Lawn, R. M., et al, ProcNatl Acad Sci (USA), 78:5435-5439 (1981)), the DNA sequence of this genedemonstrates that it lacks introns. Homology to protein sequenceinformation from these published IFN-α genes made it possible todetermine the correct translational reading frame and thus allowed theentire 166 amino acid sequence of IFN-α54 to be predicted from the DNAsequence as well as a precursor segment, or signal peptide, of 23 aminoacids (FIG. 5). The DNA sequence of the IFN-α54 gene and the amino acidsequence predicted therefrom differ substantially from the other knownIFN-α DNA and IFN-α amino acid sequences.

Plasmid Preparatiqn and Host Transformation

Assembly of the plasmid for direct expression of the IFN-α54 geneinvolved replacing the DNA fragment encoding the 23 amino acid signalpolypeptide of preinterferon with a 120 bp EcoRI/Sau3A promoter fragment(E.coli trp promoter, operator, and trp leader ribosome binding sitepreceding an ATG initiation codon) and using the naturally occurringHincII site, 139 bp 3'- of the TAA translational stop codon, to insertthe gene into a cloning vehicle derived from the plasmid pBR322. Thecomplete DNA sequence of the promoter and gene fragments insertedbetween the EcoRI and PvuII sites of pBR322 is shown in FIG. 6 whichalso shows the exact location of relevant cloning sites. Details of theconstruction are described below.

The coding region for mature IFN-α54 encompasses a Sau3A site betweencodons for amino acids 2 and 3 and an XbaI site between codons for aminoacids 81 and 83. The 239 bp Sau3A to XbaI fragment was isolated on a 6%polyacrylamide gel following a Sau3A/XbaI double-digest of the 3.9 kbEcoRI genomic fragment. This fragment was ligated to the 120 bpEcoRI/Sau 3A promoter fragment. The promoter fragment contained asynthetic HindIII restriction site, ATG initiation codon, the initialcysteine codon (TGT) and a Sau 3A "sticky end". The ligation mixture wasdigested with EcoRI and XbaI to enrich for the desired product andligated with an EcoRI/XbaI digested vector fragment pBWll (derived frompBR322 which contained unique EcoRI and XbaI restriction sites). Theligation mixture was used to transform E.coli MM294 (Backman, K., et al,Proc Natl Acad Sci (USA), 73:4174-4178 (1976)). The desired correcttransformation product, designated pCSlO, was identified by restrictionenzyme mapping. DNA from this intermediate plasmid was prepared,digested with XbaI and PvuII, and the large fragment containing thepromoter and the 5'-portion of the gene was used as a vector forreconstituting the 3'-end of the gene. Referring to the restrictionenzyme sites shown in FIG. 6, the 397 base pair Xba to HincII fragmentencompassing the 3'-codon region, the TAA translational stop codon andan additional 139 base pairs of 3'-noncoding sequence was isolated on a4% polyacrylamide gel following an XbaI/HincII double-digest of the 3.9kb genomic EcoRI fragment. This XbaI to HincII fragment was ligated tothe Xba/PvuII digested pCSlO, the ligation mixture was used to transformE.coli MM294, and correct transformants (3 out of 100 screened) wereidentified by restriction enzyme mapping. FIG. 7 is a diagram of one ofthe final expression constructs obtained, which is designated pCS12.Other prokaryotic hosts such as bacteria other than E.coli may, ofcourse, be transformed with this or other suitable constructs toreplicate the IFN-α54 gene and/or to produce IFN-α54.

Cultivation of Transformants

Bacteria transformed with the IFN-α54 gene may be cultivated in anappropriate growth medium, such as a minimum essential medium, thatsatisfies the nutritional and other requirements needed to permit thebacteria to grow and produce IFN-α54. If the bacteria are such that theprotein is contained in their cytoplasm, the IFN-α54 may be extractedfrom the cells by lysing the cells such as by sonication and/ortreatment with a strong anionic solubilizing agent such as sodiumdodecyl sulfate. Further purification of the extract may be achieved byaffinity chromatography, electrophoresis, or other protein purificationtechniques.

IFN-α54 produced in accordance with the invention is believed to bedistinct from the corresponding native protein in several respects.Firstly, because the IFN-α54 gene was expressed by bacterial hosts thatutilize N-formyl-methionine and/or methionine to initiate translation,some or all of the bacterially produced IFN-α54 molecules are precededby an N-formyl-methionine or methionine group. Some of theN-formyl-methionine or methionine groups could be removed by natural invivo bacterial cleavage mechanisms. This would result in a mixture ofmolecules, some of which would include an initial N-formyl-methionine ormethionine and others that would not. All such IFN-α54 molecules, thosecontaining an initial N-formyl-methionine or methionine, those notcontaining an N-formyl-methionine or methionine and any mixture thereof,are encompassed by the present invention. Secondly, the amino acidresidues of the bacterially produced polypeptide are unsubstitutedwhereas the residues of the native protein may be substituted with sugargroups, ACTH or other moieties. Also, native IFN-α extracts consist ofmixtures of various IFN molecules whereas the bacterially producedIFN-α54 is homogeneous; that is, bacterially produced IFN-α54 does notcontain functionally related polypeptides. Accordingly, the inventioncontemplates producing IFN-α54-containing compositions having biologicalactivity that is attributable solely to IFN-α54 and/or said terminalN-formyl-methionine or methionine derivatives thereof.

Biological Testing of IFN-α54

IFN-α54-containing cell sonicates were tested in vitro and found to havethe following activities: (1) inhibition of viral replication ofvesicular stomatitis virus (VSV) and herpes simplex virus-1 (HSV-1); (2)inhibition of tumor cell growth; (3) inhibition of colony formation bytumor cells in soft agar; (4) activation of natural killer (NK) cells;(5) enhancement of the level of 2',5'-oligoadenylate synthetase(2',5'-A); and (6) enhancement of the double-stranded RNA-dependentprotein kinase. The sonicates were active in inhibiting viral infectionin both human and other mammalian cells such as hamster, monkey, mouse,and rabbit cells.

The tests show that IFN-α54 exhibits antiviral activity against DNA andRNA viruses, cell growth regulating activity, and an ability to regulatethe production of intracellular enzymes and other cell-producedsubstances. Accordingly, it is expected IFN-α54 may be used to treatviral infections with a potential for interferon therapy such as chronichepatitis B infection, ocular, local, or systemic herpes virusinfections, influenza and other respiratory tract virus infections,rabies and other viral zoonoses, arbovirus infections, and slow virusdiseases such as Kuru and sclerosing panencephalitis. It may also beuseful for treating viral infections in immunocompromised patients suchas herpes zoster and varicella, cytomegalovirus, Epstein-Barr virusinfection, herpes simplex infections, rubella, and progressivemultifocal leukoencephalopathy. Its cell growth regulating activitymakes it potentially useful for treating tumors and cancers such asosteogenic sarcoma, multiple myeloma, Hodgkin's disease, nodular, poorlydifferentiated lymphoma, acute lymphocytic leukemia, breast carcinoma,melanoma, and nasopharyngeal carcinoma. The fact that IFN-α54 increasesprotein kinase and 2',5'-oligoadenylate synthetase indicates it may alsoincrease synthesis of other enzymes or cell-produced substances commonlyaffected by IFNs such as histamine, hyaluronic acid, prostaglandin E,tRNA methylase, and aryl hydrocarbon hydrolase. Similarly, it may beuseful to inhibit enzymes commonly inhibited by IFNs such as tyrosineamino transferase, glycerol-3-phosphate dehydrogenase glutaminesynthetase, ornithine decarboxylase, S-adenosyl-1-methioninedecarboxylase, and UDP-N-acetylglucosamine-dolichol monophosphatetransferase. The ability of the IFN-α54 to stimulate NK cell activity isindicative that it may also possess other activities such as theabilities to induce macrophage activity and antibody production and toeffect cell surface alterations such as changes in plasma membranedensity or cell surface charge, altered capacity to bind substances suchas cholera toxin, concanavalin A and thyroid-stimulating hormone, andchange in the exposure of surface gangliosides.

Pharmaceutical compositions that contain IFN-α54 as an active ingredientwill normally be formulated with an appropriate solid or liquid carrierdepending upon the particular mode of administration being used. Forinstance, parenteral formulations are usually injectable fluids that usepharmaceutically and physiologically acceptable fluids such asphysiological saline, balanced salt solutions, or the like as a vehicle.Oral formulations, on the other hand, may be solid, e.g. tablet orcapsule, or liquid solutions or suspensions. IFN-α54 will usually beformulated as a unit dosage form that contains in the range of 10⁴ to10⁷ international units, more usually 10⁶ to 10⁷ international units,per dose.

IFN-α54 may be administered to humans in various manners such as orally,intravenously, intramuscularly, intraperitoneally, intranasally,intradermally, and subcutaneously. The particular mode of administrationand dosage regimen will be selected by the attending physician takinginto account the particulars of the patient, the disease and the diseasestate involved. For instance, viral infections are usually treated bydaily or twice daily doses over a few days to a few weeks; whereas tumoror cancer treatment involves daily or multidaily doses over months oryears. IFN-α54 therapy may be combined with other treatments and may becombined with or used in association with other chemotherapeutic orchemopreventive agents for providing therapy against viral infections,neoplasms, or other conditions against which it is effective. Forinstance, in the case of herpes virus keratitis treatment, therapy withIFN has been supplemented by thermocautery, debridement andtrifluorothymidine therapy.

Modifications of the above described modes for carrying out theinvention, such as, without limitation, use of alternative vectors,alternative expression control systems in the vector, and alternativehost microorganisms and other therapeutic or related uses of IFN-α54,that are obvious to those of ordinary skill in the biotechnology,pharmaceutical, medical and/or related fields are intended to be withinthe scope of the following claims.

I claim:
 1. A recombinantly produced polypeptide having interferonactivity and comprising the amino acid sequence of IFN-α54, :

    ______________________________________                                        CysAspLeuProGln    ThrHisSerLeuGly                                            HisArgArgThrMet    MetLeuLeuAlaGln                                            MetArgArgIleSer    LeuPheSerCysLeu                                            LysAspArgHisAsp    PheArgPheProGln                                            GluGluPheAspGly    AsnGlnPheGlnLys                                            AlaGluAlaIleSer    ValLeuHisGluVal                                            IleGlnGlnThrPhe    AsnLeuPheSerThr                                            LysAspSerSerVal    AlaTrpAspGluArg                                            LeuLeuAspLysLeu    TyrThrGluLeuTyr                                            GlnGlnLeuAsnAsp    LeuGluAlaCysVal                                            MetGlnGluValTrp    ValGlyGlyThrPro                                            LeuMetAsnGluAsp    SerIleLeuAlaVal                                            ArgLysTyrPheGln    ArgIleThrLeuTyr                                            LeuThrGluLysLys    TyrSerProCysAla                                            TrpGluValValArg    AlaGluIleMetArg                                            SerPheSerSerSer    ArgAsnLeuGlnGlu                                            ArgLeuArgArgLys    Glu.                                                       ______________________________________                                    


2. The polypeptide of claim 1 wherein the polypeptide consistsessentially of said amino acid sequence.
 3. The polypeptide of claim 1or 2 wherein the initial cysteine residue of the amino acid sequence ispreceded by an N-formyl-methionine group.
 4. The polypeptide of claim 1or 2 wherein the amino acid residues of said sequence are unsubstituted.5. Recombinantly produced IFN-α54.
 6. A composition having interferonactivity and comprising a mixture of:(a) a precombinantly producedpolypeptide having the amino acid sequence

    ______________________________________                                        CysAspLeuProGln    ThrHisSerLeuGly                                            HisArgArgThrMet    MetLeuLeuAlaGln                                            MetArgArgIleSer    LeuPheSerCysLeu                                            LysAspArgHisAsp    PheArgPheProGln                                            GluGluPheAspGly    AsnGlnPheGlnLys                                            AlaGluAlaIleSer    ValLeuHisGluVal                                            IleGlnGlnThrPhe    AsnLeuPheSerThr                                            LysAspSerSerVal    AlaTrpAspGluArg                                            LeuLeuAspLysLeu    TyrThrGluLeuTyr                                            GlnGlnLeuAsnAsp    LeuGluAlaCysVal                                            MetGlnGluValTrp    ValGlyGlyThrPro                                            LeuMetAsnGluAsp    SerIleLeuAlaVal                                            ArgLysTyrPheGln    ArgIleThrLeuTyr                                            LeuThrGluLysLys    TyrSerProCysAla                                            TrpGluValValArg    AlaGluIleMetArg                                            SerPheSerSerSer    ArgAsnLeuGlnGlu                                            ArgLeuArgArgLys    Glu                                                        ______________________________________                                    

and; (b) a polypeptide having said amino acid sequence wherein theinitial cysteine residue of the sequence is preceded by anN-formyl-methionine or methionine group.
 7. The composition of claim 6wherein the amino acid residues of said sequence are unsubstituted.
 8. Acomposition having interferon activity comprising a polypeptide havingthe amino acid sequence

    ______________________________________                                        CysAspLeuProGln    ThrHisSerLeuGly                                            HisArgArgThrMet    MetLeuLeuAlaGln                                            MetArgArgIleSer    LeuPheSerCysLeu                                            LysAspArgHisAsp    PheArgPheProGln                                            GluGluPheAspGly    AsnGlnPheGlnLys                                            AlaGluAlaIleSer    ValLeuHisGluVal                                            IleGlnGlnThrPhe    AsnLeuPheSerThr                                            LysAspSerSerVal    AlaTrpAspGluArg                                            LeuLeuAspLysLeu    TyrThrGluLeuTyr                                            GlnGlnLeuAsnAsp    LeuGluAlaCysVal                                            MetGlnGluValTrp    ValGlyGlyThrPro                                            LeuMetAsnGluAsp    SerIleLeuAlaVal                                            ArgLysTyrPheGln    ArgIleThrLeuTyr                                            LeuThrGluLysLys    TyrSerProCysAla                                            TrpGluValValArg    AlaGluIleMetArg                                            SerPheSerSerSer    ArgAsnLeuGlnGlu                                            ArgLeuArgArgLys    Glu                                                        ______________________________________                                    

or a mixture of said polypeptide and a polypeptide having said sequencewherein the initial cysteine residue is preceded by anN-formyl-methionine or methionine group wherein the interferon activityof the composition is attributable to said polypeptide or to saidmixture.
 9. A pharmaceutical composition for providing interferontherapy to a mammal comprising an effective amount of the polypeptide ofclaim 1, 2, or 5 admixed with a pharmaceutically acceptable vehicle orcarrier.
 10. A pharmaceutical composition for providing interferontherapy to a mammal comprising an effective amount of the composition ofclaims 6 or 8 admixed with a pharmaceutically acceptable vehicle orcarrier.
 11. A method of providing interferon therapy to a humancomprising administering a therapeutically effective amount of thepolypeptide of claim 1, 2 or 5 to said human.
 12. A method of providinginterferon therapy to a human comprising administering a therapeuticallyeffective amount of the composition of claim 6 or 8 to said human. 13.The method of claim 12 wherein the therapy is for treating a viralinfection, providing cell growth regulation, or regulating theproduction of a cell-produced substance.
 14. The method of claim 13wherein the therapy is for treating a viral infection, providing cellgrowth regulation, or regulating the production of a cell-producedsubstance.
 15. A method of providing antiviral therapy to a mammalcomprising administering a viral infection inhibiting amount of thepolypeptide of claim 1, 2 or 5 to the mammal.